CN105104181A - Haploid genome doubling method for maize - Google Patents
Haploid genome doubling method for maize Download PDFInfo
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- CN105104181A CN105104181A CN201510503664.4A CN201510503664A CN105104181A CN 105104181 A CN105104181 A CN 105104181A CN 201510503664 A CN201510503664 A CN 201510503664A CN 105104181 A CN105104181 A CN 105104181A
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Abstract
The invention belongs to the field of agricultural maize breeding and particularly relates to a haploid genome doubling method for maize. The haploid genome doubling method for the maize comprises the following steps: (1) carrying out hybridization in a manner of taking agricultural university highly-induced 1# as a male parent and taking a CML495*PLS hybridized combination as a female parent, so as to obtain maize haploid seeds; (2) sowing the maize haploid seeds into a soilless culture medium, so as to obtain maize haplobiont seedlings; (3) when the maize haplobiont seedlings grow to a 2-3-leaf stage, changing a nutrient solution for soilless culture into a colchicine solution, carrying out culturing for 8-10 hours, and then, changing the colchicine solution into the nutrient solution for soilless culture; and (4) transplanting the haploid maize seedlings treated with the colchicine solution to fields, and harvesting seeds, i.e., maize diploid seeds. The method disclosed by the invention has the advantages of simplicity in operation, high doubling rate and high human safety, the workload for scientific research personnel during maize breeding is reduced, and the time for maize breeding is shortened.
Description
Technical field
The invention belongs to agriculture corn breeding field, particularly relate to a kind of method that corn haploid chromosomes group doubles.
Background technology
Owing to only overlapping chromosome set containing 1, the general growing way of corn haplobiont is more weak, and plant is short and small, often shows highly sterile, is difficult to directly apply in production.Exterior technology is utilized the chromosome doubling of monoploid individuality to be allowed its loose powder of normally weaving silk obtain.Corn doubling monoploids technology is the bottleneck of current corn haploid breeding technology application.
Double haploid (Doubledhaploid, DH) breeding technique induction Haploid production, again through doubling to form dliploid, general only need through 2 seasons of growth just can obtain 100% pure lines, this technology is called as DH (DoubledHaploid) technology, DH technology can accelerate breeding process, is the normal technology adopted in modern corn breeding.
At present, China breeder mainly adopts two kinds of approach to carry out haploid chromosomes to double: monoploid Natural double or monoploid artificial doubling.
Monoploid Natural double, due to not by any medicament, is the method for less expensive material benefit.But Natural double is only applicable to the high background material of Natural doubling rate, actively will create geography, ecology, the weather conditions that can improve monoploid Natural doubling rate, the method affects comparatively large by genetic background and to double success rate low simultaneously.
Monoploid artificial doubling obtains DH system comparatively effective method, the most frequently used is chemical method, its principle is the formation of spindle fiber when using chemical agent destruction cell mitogen, 2 sub-cell chromosomes are made not shift to the two poles of the earth in mitosis anaphase, thus change cell chromosome ploidy, realize chromosomal chemistry and double.At present, the main chemical agent utilized has colchicine and weed killer herbicide.Chemical method is divided into again infusion method and drip: infusion method is generally the colchicine solution soaking corn seedling apical point with 0.06-0.5%, soak time is 12-48h, proceed to hot-house culture or move into field plantation after again material clean is clean, adding multiplying power can reach 20-50%.Be widely used because the method doubling etticiency is higher always, but the method needs to use the long-time soaking corn seedling of colchicine solution, then carry out cleaning and transplanting, high malicious carcinogenic substance-colchicine is in open environment for a long time, and the healthy safety of scientific research personnel is often on the hazard; Drip instils in cotyledon or seedling apical point with dropper by the colchicine solution of 0.06-0.5%, need every 6-8h titration repeatedly, program is loaded down with trivial details, and workload is large, and staff is exposed in the open environment of colchicine in titration process repeatedly, to be healthyly on the hazard.
Therefore, modern corn breeding field be badly in need of a kind of success rate high, again can the healthy corn haploid chromosomes group method for doubling of safeguard work personnel.
Summary of the invention
The object of the present invention is to provide one to improve and double success rate, the method that the corn haploid chromosomes group preserved to the health of scientific research personnel again doubles.
For achieving the above object, technical scheme provided by the invention is as follows:
The method that corn haploid chromosomes group doubles, is characterized in that, comprise the following steps:
(1) lure No. 1 for male parent so that agricultural university is high, be maternal with CML495 × PLS hybrid combination, hybridization obtains corn monoploid seed;
(2) step (1) gained corn monoploid planting seed is cultivated in soilless culture medium, make its rudiment, obtain corn haplobiont seedling;
(3) when corn haplobiont growth of seedling is to the 2-3 leaf phase, change the nutrient solution of soilless culture into colchicine solution, incubation time is 8-10h, and then colchicine solution is changed into the nutrient solution of soilless culture;
(4) then move to land for growing field crops by with the monoploid corn seedling of colchicine solution process, results seed, is corn dliploid seed.
In technical scheme of the present invention, the matrix of the soilless culture medium described in step (2) is vermiculite, and in described soilless culture medium, contained nutrition percentage is: N5.12%, P2.72% and K7.04%.
As preferably, the corn haplobiont seedling described in step (2) is that corn monoploid seed is sowed and obtained in green house.
In technical scheme of the present invention, the mass concentration of the colchicine solution described in step (3) is 0.025-0.125%.
As preferably, the mass concentration of described colchicine solution is 0.075%.
As preferably, the incubation time described in step (3) is 9h.
Compared with prior art, the present invention has following beneficial effect:
(1) method of the present invention is passed through soilless culture, is used colchicine soaking corn seedling root, the effect of colchicine is made to reach maximization, double success rate and can bring up to 35-55%, higher than using, the 1-5%'s of Natural double method add multiplying power, reduce the workload of scientific research personnel, shorten breeding time.
(2) method of the present invention is simpler than traditional chemical medical method, quick, scientific research personnel carries out soilless culture after only needing configuration colchicine solution, and program is simple, and scientific research personnel is without the need to being exposed in the environment of colchicine pollution, health does not suffer damage, and is healthy and safely protected.
Embodiment
Following examples for illustration of the present invention, but are not used for limiting the scope of the invention.When not deviating from the present invention's spirit and essence, the amendment do the inventive method, step or condition or replacement, all belong to scope of the present invention.If do not specialize, chemical reagent used in embodiment, seed, seedling are conventional commercial, the conventional means that technological means used in embodiment is well known to those skilled in the art.
CML495 and PLS in embodiment is the corn inbred line of international corn wheat improvement center (CIMMYT) of Yin Zi, and the multiple diseases such as CML495 high-resistance corn northern and southern leaf blight and QTL mapping, have higher general combining ability; The resistance to low nitrogen of PLS has higher general combining ability simultaneously, and both filial generations should be easier to seed selection and obtain high-combining ability and the corn inbred line adapting to Guangxi ecological condition.
embodiment1:
In March, (1) 2013 lures No. 1 for male parent so that agricultural university is high, two CML495 × PLS hybrid combination is maternal, hybridization acquisition 126 corn monoploid seeds, in August, 2013 by these monoploid planting seeds in corn research institute of Guangxi academy of agricultural sciences green house, carry out soilless culture, make its sprout growth;
(2) configuration quality percentage is the colchicine solution of 0.025%, treat corn germination and grow into 3 leaf full leaf stretching period micro pump the nutrient solution of soilless culture is changed into the colchicine solution of 0.025% and cultivates 8h, finally again the colchicine solution of 0.025% is changed into the nutrient solution of soilless culture;
(3) move to land for growing field crops and meticulous nursing with the monoploid corn seedling after colchicine solution process, results seed, is amphiploid corn seed.
embodiment 2:
In March, (1) 2013 lures No. 1 for male parent so that agricultural university is high, two CML495 × PLS hybrid combination is maternal, hybridization acquisition 110 corn monoploid seeds, in August, 2013 by these planting seeds in corn research institute of Guangxi academy of agricultural sciences green house, carry out soilless culture, make its sprout growth;
(2) configuration quality percentage is the colchicine solution of 0.075%, treat corn germination and grow into 3 leaf full leaf stretching period micro pump the nutrient solution of soilless culture is changed into the colchicine solution of 0.075% and cultivates 9h, finally the colchicine solution of 0.075% is changed into the nutrient solution of soilless culture;
(3) land for growing field crops is moved to and meticulous nursing with the monoploid corn seedling after colchicine solution process, results seed.
embodiment 3:
In March, (1) 2013 lures No. 1 for male parent so that agricultural university is high, two CML495 × PLS hybrid combination is maternal, hybridization acquisition 132 corn monoploid seeds, in August, 2013 by these planting seeds in corn research institute of Guangxi academy of agricultural sciences green house, carry out soilless culture, make its sprout growth;
(2) configuration quality percentage is the colchicine solution of 0.125%, treat corn germination and grow into 3 leaf full leaf stretching period micro pump the nutrient solution of soilless culture is changed into the colchicine solution of 0.125% and cultivates 10h, finally again the colchicine solution of 0.125% is changed into the nutrient solution of soilless culture;
(3) land for growing field crops is moved to and meticulous nursing with the monoploid corn seedling after colchicine solution process, results seed.
result of implementation and analysis:
Through statistics, the result of implementation of embodiment 1-3 is respectively (referring to table 1):
Embodiment 1:2013 has 58 strain plant loose powder October, and namely doubling success rate is 58/126 × 100%=46.0%, and in time gathering in the crops by the end of November, totally 45 strain corns can receive at least 1 seed, and solid strain rate is 45/126=35.7%; Sow this batch of seeds totally 45 head progeny rows in March, 2014, carry out Observation on Agronomic Characters and combining ability test, be finally bred as the double haploid corn inbred line that 16 economical characters are excellent.If use Natural double method to carry out chromosome set to double, need the corn monoploid seed of about 1933, mean and need CML495 × PLS filial generation of hybridization 15.3 times to be that female parent is induced and just can be reached similar effect.
Embodiment 2:2013 has 58 strain plant loose powder October, and namely doubling success rate is 58/110 × 100%=52.7%, and in time gathering in the crops by the end of November, totally 45 strain corns can receive at least 1 seed, and solid strain rate is 45/110=40.9%; Sow this batch of seeds totally 45 head progeny rows in March, 2014, carry out Observation on Agronomic Characters and combining ability test, be finally bred as the double haploid corn inbred line that 16 economical characters are excellent.If use Natural double method to carry out chromosome set to double, need the corn monoploid seed of about 1933, mean and need CML495 × PLS filial generation of hybridization 17.6 times to be that female parent is induced and just can be reached similar effect.
Embodiment 3:2013 has 58 strain plant loose powder October, and namely doubling success rate is 58/132 × 100%=52.7%, and in time gathering in the crops by the end of November, totally 45 strain corns can receive at least 1 seed, and solid strain rate is 45/132=40.9%; Sow this batch of seeds totally 45 head progeny rows in March, 2014, carry out Observation on Agronomic Characters and combining ability test, be finally bred as the double haploid corn inbred line that 16 economical characters are excellent.If use Natural double method to carry out chromosome set to double, need the corn monoploid seed of about 1933, mean and need CML495 × PLS filial generation of hybridization 14.6 times to be that female parent is induced and just can be reached similar effect.
Table 1 the inventive method and Natural double method doubling etticiency contrast
In sum, method of the present invention is used to double corn Chromosomes of Haploid group, double success rate and reach 52.7%, solid strain rate can reach 40.9%, far above the solid strain rate using the 1-5% of Natural double method to add multiplying power and 0.5-8%, visible, the inventive method can be increased work efficiency greatly, and scientific research personnel is also without the need to being exposed to the environment that carcinogenic substance colchicine pollutes, high to human safety.
The aforementioned description to concrete exemplary of the present invention is to illustrate and the object of illustration.These descriptions not want the present invention to be defined as disclosed precise forms, and obviously, according to above-mentioned instruction, can much change and change.The object selected exemplary embodiment and describe is to explain certain principles of the present invention and practical application thereof, thus those skilled in the art can be realized and utilize various different exemplary of the present invention and various different selection and change.Scope of the present invention is intended to limited by claims and equivalents thereof.
Claims (6)
1. the method that doubles of corn haploid chromosomes group, is characterized in that, comprise the following steps:
(1) lure No. 1 for male parent so that agricultural university is high, be maternal with CML495 × PLS hybrid combination, hybridization obtains corn monoploid seed;
(2) step (1) gained corn monoploid planting seed is cultivated in soilless culture medium, make its rudiment, obtain corn haplobiont seedling;
(3) when corn haplobiont growth of seedling is to the 2-3 leaf phase, change the nutrient solution of soilless culture into colchicine solution, incubation time is 8-10h, and then colchicine solution is changed into the nutrient solution of soilless culture;
(4) then move to land for growing field crops by with the monoploid corn seedling of colchicine solution process, results seed, is corn dliploid seed.
2. the method that doubles of corn haploid chromosomes group according to claim 1, it is characterized in that, the matrix of the soilless culture medium described in step (2) is vermiculite, and in described soilless culture medium, contained nutrition percentage is: N5.12%, P2.72% and K7.04%.
3. the method that doubles of corn haploid chromosomes group according to claim 1, is characterized in that, the corn haplobiont seedling described in step (2) is that corn monoploid seed is sowed and obtained in green house.
4. the method that doubles of corn haploid chromosomes group according to claim 1, it is characterized in that, the mass concentration of the colchicine solution described in step (3) is 0.025-0.125%.
5. the method that doubles of corn haploid chromosomes group according to claim 4, it is characterized in that, the mass concentration of described colchicine solution is 0.075%.
6. the method that doubles of corn haploid chromosomes group according to claim 1, it is characterized in that, the incubation time described in step (3) is 9h.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109392700A (en) * | 2018-12-17 | 2019-03-01 | 靖西市秀美边城农业科技有限公司 | A kind of method that corn × rice distant hybridization is bred as dihaploid |
| CN111713403A (en) * | 2020-07-31 | 2020-09-29 | 金苑(北京)农业技术研究院有限公司 | Method for doubling corn haploid seedlings |
| CN112568077A (en) * | 2020-12-14 | 2021-03-30 | 吉林省农业科学院 | Method for improving doubling efficiency of corn haploid |
-
2015
- 2015-08-17 CN CN201510503664.4A patent/CN105104181A/en active Pending
Non-Patent Citations (1)
| Title |
|---|
| 刘鹏宇等: "玉米活体诱导单倍体的加倍方法研究进展", 《安徽农业科学》 * |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109392700A (en) * | 2018-12-17 | 2019-03-01 | 靖西市秀美边城农业科技有限公司 | A kind of method that corn × rice distant hybridization is bred as dihaploid |
| CN111713403A (en) * | 2020-07-31 | 2020-09-29 | 金苑(北京)农业技术研究院有限公司 | Method for doubling corn haploid seedlings |
| CN111713403B (en) * | 2020-07-31 | 2022-02-18 | 金苑(北京)农业技术研究院有限公司 | Method for doubling corn haploid seedlings |
| CN112568077A (en) * | 2020-12-14 | 2021-03-30 | 吉林省农业科学院 | Method for improving doubling efficiency of corn haploid |
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